Optogenetics is the combination of genetic and optical methods used to control specific events in targeted cells of living tissue, even within freely moving mammals and other animals, with the temporal precision (millisecond-timescale) needed to keep pace with functioning intact biological systems. The hallmark of optogenetics is the introduction of fast light-activated channel proteins to the plasma membranes of target neuronal cells that allow temporally precise manipulation of neuronal membrane potential while maintaining cell-type resolution through the use of specific targeting mechanisms. Among the microbial opsins which can be used to investigate the function of neural systems are the channelrhodopsins (ChR2, ChR1, VChR1, and SFOs) used to promote depolarization in response to light. In just a few short years, the field of optogenetics has furthered the fundamental scientific understanding of how specific cell types contribute to the function of biological tissues such as neural circuits in vivo. Moreover, on the clinical side, optogenetics-driven research has led to insights into Parkinson's disease and other neurological and psychiatric disorders.
However, in spite of these advances, the neurophysiological substrates of most psychiatric disorders remain poorly understood, despite rapidly emerging information on genetic factors that are associated with complex behavioral phenotypes such as those observed in autism and schizophrenia (Cichonet al., The American Journal of Psychiatry 166(5):540 (2009); O'Donovan et al., Human Genetics 126(1): 3 (2009)). One remarkable emerging principle is that a very broad range of seemingly unrelated genetic abnormalities can give rise to the same class of psychiatric phenotype (such as social behavior dysfunction; Folstein & Rosen-Sheidley, Nature Reviews 2(12):943 (2001)). This surprising pattern has pointed to the need to identify simplifying circuit-level insights that could unify diverse genetic factors under a common pathophysiological principle.
One such circuit-level hypothesis is that elevation in the ratio of cortical cellular excitation and inhibition (cellular E/I balance) could give rise to the social and cognitive deficits of autism (Rubenstein, Current Opinion in Neurology 23(2):118; Rubenstein & Merzenich, Genes, Brain, and Behavior 2(5):255 (2003)). This hypothesis could potentially unify diverse streams of pathophysiological evidence, including the observation that many autism-related genes are linked to gain-of-function phenotypes in ion channels and synaptic proteins (Bourgeron, Current Opinion in Neurobiology 19 (2), 231 (2009)) and that ˜30% of autistic patients also show clinically apparent seizures (Gillberg & Billstedt, Acta Psychiatrica Scandinavica, 102(5):321 (2000)). However, it has not been clear if such an imbalance (to be relevant to disease symptoms) would be operative on the chronic (e.g. during development) or the acute timescale. Furthermore, this hypothesis is by no means universally accepted, in part because it has not yet been susceptible to direct testing. Pharmacological and electrical interventions lack the necessary specificity to selectively favor activity (in a manner fundamentally distinct from receptor modulation) of neocortical excitatory cells over inhibitory cells, whether in the clinical setting or in freely behaving experimental mammals during social and cognitive tasks. It is perhaps related to challenges such as this that the social and cognitive deficits of autism and schizophrenia have proven largely unresponsive to conventional psychopharmacology treatments in the clinic.
Existing optogenetic methods are also inadequate for this purpose; driving coordinated spikes selectively in excitatory or inhibitory cells with a channelrhodopsin is feasible, but not well suited to the sparse coding and asynchronous firing patterns of neocortical pyramidal cells. Moreover, the continuous presence of an optical fiber and other hardware poses challenges for prolonged behavioral tests with fast and spatially complex movements typical of social behavior and cognitive measures (for example in contextual conditioning). Instead, selectively favoring excitation of one population over another with a bistable step-function opsin (SFO) gene product could partially address these challenges, since the targeted population would not be driven with coordinated spikes, but merely sensitized to native inputs that can be sparse and asynchronous. Use of SFOs also has the potential to address the hardware challenge, since the orders-of-magnitude greater light sensitivity characteristic of SFOs could in theory allow non-brain penetrating light delivery, and the persistent action of the bistable SFOs after light-off could allow hardware-free behavioral testing. However, the known SFOs (C128A,S,T and D156A) are not stable enough to produce constant photocurrent after a single light flash over the many minutes required for complex behavioral testing.
What is needed, therefore, is an optogenetic tool which would permit direct testing of the E/I balance hypothesis in the prefrontal cortex both in vitro and in vivo in freely-moving mice. Such a light-activated protein could permit investigation of the effect of bi-directional modulation of prefrontal cellular E/I balance on both conditioned and innate behaviors relevant for cognitive and social dysfunction, as well as probe the resulting effects on circuit physiology and quantitative transmission of information.